Developmental strategy of the endoparasite Xenos vesparum (strepsiptera, Insecta): host invasion and elusion of its defense reactions

To successfully complete its endoparasitic development, the strepsipteran Xenos vesparum needs to elude the defense mechanisms of its host, the wasp Polistes dominulus. SEM and TEM observations after artificial infections allow us to outline the steps of this intimate host-parasite association. Triungulins, the mobile 1st instar larvae of this parasite, are able to "softly" overcome structural barriers of the larval wasp (cuticle and epidermis) without any traumatic reaction at the entry site, to reach the hemocoel where they settle. The parasite molts 48 h later to a 2nd instar larva, which moves away from the 1st instar exuvium, molts twice more without ecdysis (a feature unique to Strepsiptera) and pupates, if male, or develops into a neotenic female. Host encapsulation involves the abandoned 1st larval exuvium, but not the living parasite. In contrast to the usual process of encapsulation, it occurs only 48 h after host invasion or later, and without any melanization. In further experiments, first, we verified Xenos vesparum's ability to reinfect an already parasitized wasp larva. Second, 2nd instar larvae implanted in a new host did not evoke any response by hemocytes. Third, we tested the efficiency of host defense mechanisms by implanting nylon filaments in control larval wasps, excluding any effect due the dynamic behavior of a living parasite; within a few minutes, we observed the beginning of a typical melanotic encapsulation plus an initial melanization in the wound site. We conclude that the immune response of the wasp is manipulated by the parasite, which is able to delay and redirect encapsulation towards a pseudo-target, the exuvia of triungulins, and to elude hemocyte attack through an active suppression of the immune defense and/or a passive avoidance of encapsulation by peculiar surface chemical properties.     

Factors influencing the developmental capacity of Galleria larval epidermis

The nature of the cuticle secreted by integument from a day-1 penultimate instar larval Galleria when cultured in vivo in the abdomen of a last instar larva varied with the age of the host. When placed in a day-5 last instar larva, the implanted integument secreted a pupal cuticle at the time the host metamorphosed and became a pupa. However, when placed in a day-7 last instar larva the implant, from the same stage donor, secreted a larval cuticle at the time the host pupated. Experimental studies involving implantation of the integument for a 24 hr period, into various developmental stages of normal and ligated last instar larvae, pupae, and pharate adults, prior to placing it in a day-7 last instar larva suggest that a non-hormonal factor present in day-4 and -5 last instar larvae is important to initiate pupal syntheses.     

Reprogramming in the absence of DNA synthesis in Galleria larval epidermis.

Larval epidermal cells from a day-1 penultimate instar Galleria larva on implantation into day-5 last instar larva metamorphose and deposit a pupal cuticle at the same time as the host pupates. DNA synthesis in the implanted larval cell was monitored with 3-H-thymidine. Various regimens of 3-H-thymidine application were used and under no conditions did the larval cells incorporate label during the period from implantation to deposition of pupal cuticle. This suggests that a wax moth larval ectoderm cell can reprogram its genome to secrete a pupal cuticle without a precedent cell division.     

A post-ecdysial plasticization of the abdominal cuticle in Rhodnius

The ecdysis and emergence of fifth instar larvae of Rhodnius prolixus have been closely observed. At the time of ecdysis the cuticle of the head, legs, and wingpads is soft and readily deformed. it does not become sufficiently rigid for normal use until about 90 min later. The cuticle of the abdomen is however hard and inextensible at the time of ecdysis. From about 60 min onward this cuticle undergoes a plasticization; it is maximally extensible at about 180 min, thereafter becoming inextensible again. Unlike the plasticization of the abdominal cuticle which occurs after feeding, this post-ecdysial plasticization is not under direct nervous control. Although it seems that there is some temporal link with the darkening of the cuticle, it is considered unlikely that plasticization is a direct consequence of the tanning process. The significance of this post-ecdysial plasticization is not obvious.     

Correlations between epidermal cell structure and endogenous hormone titers during the fifth larval instar of the tobacco hornworm, Manduca sexta

Epidermal cell morphology and cuticle production in Manduca sexta are directly influenced by both ecdysterone and juvenile hormone. Up to day 6 of the last larval instar, post-molt endocuticle is continuously deposited even though cells undergo a partial and temporary separation from the overlying cuticle at the time when a small ecdysteroid peak is detected (approximately day 3.5). At about days 6--7 when another, larger ecdysteroid peak is present, apolysis occurs accompanied by the appearance of edcysial droplets. Following apolysis, layers of pupal cuticle are deposited. Increased quantities of rough endoplasmic reticulum characterize the epidermis at times of peak endocuticle deposition (day 3, larval cuticle; day 9, pupal cuticle). Dense pigment inclusions are found in epidermis from the day of ecdysis to the last larval instar until they are eliminated 5 days later. These dense bodies migrate from cell apex to base in the absence of juvenile hormone (or in the presence of a negligible amount of juvenile hormone) and probably contain insecticyanin.     

嘴壶夜蛾的形态、生活史及昼夜节律

嘴壶夜蛾Oraesia emarginata (Fabricius)是危害水果果实的重要害虫之一, 其成虫和幼虫取食不同的寄主植物, 可以作为理想的嗅觉研究模式昆虫。为了全面地了解嘴壶夜蛾的形态特性和生物学特性, 本实验通过室内饲喂和红外摄像机观察, 对嘴壶夜蛾各虫态的外部形态、 发育以及昼夜活动节律进行了系统研究。结果表明： 嘴壶夜蛾的各龄幼虫可以通过体色, 体表色斑的类型、 位置和数量, 以及腹足的数量进行区别。通过蛹的生殖孔和成虫触角能够很好地区别雌雄。在室内饲养条件下, 嘴壶夜蛾的寿命为53.18±1.70 d, 存活率为63.62%±2.15%, 其中幼虫的发育历期最长, 存活率最低, 卵的发育历期最短, 存活率最高, 雄成虫的存活时间显著长于雌成虫（P=0.008）。6龄幼虫的发育历期（5.29±0.15 d）显著长于其余各龄幼虫（P<0.001）。同一龄发育中期幼虫的体长和体重显著大于将蜕皮幼虫和刚蜕皮幼虫（P≤0.037）; 第2-6龄刚蜕皮幼虫的体重和体长与前一龄发育中期幼虫之间没有显著差异（P≥0.106）。幼虫在光期的孵化、 蜕皮和化蛹比例高于暗期, 而成虫在暗期的飞行、 产卵比例高于光期, 成虫的飞行随着暗期时间的增加而逐渐变少, 光期成虫的飞行主要在开灯之后1 h。成虫交配集中在暗期的第3-5 小时。本研究结果有助于制定有效的嘴壶夜蛾防治措施, 而且为嘴壶夜蛾作为嗅觉研究模式奠定基础。     

The Drosophila hugin gene codes for myostimulatory and ecdysis-modifying neuropeptides

In a genomic screen we isolated the Drosophila gene hugin (hug, cytology 87C1-2) by cross-hybridisation to a human glial cell line-derived neurotrophic factor cDNA. Upon cDNA sequence analysis and in vitro expression assays, the hugin gene was found to encode a signal peptide containing proprotein that was further processed in Schneider-2 cells into peptides similar to known neuropeptides. Two of the peptides were similar to FXPRL-amides (pyrokinins) and to the ecdysis-triggering hormone, respectively. The former displayed myostimulatory activity in a bioassay on the cockroach hyperneural muscle preparation, as well as in the Drosophila heart muscle assay. Hugin is expressed during the later half of embryogenesis and during larval stages in a subgroup of neurosecretory cells of the suboesophageal ganglion. Ubiquitous ectopic hugin expression resulted in larval death predominantly at or shortly after ecdysis from second to third instar, suggesting that at least one of the posttranslational cleavage products affects molting of the larva by interfering with the regulation of ecdysis.     

Juvenoid action on the total body metabolism in larvae of a noctuid moth

A juvenoid compound known as methoprene has no effect on growth and respiratory metabolism in penultimate instar larvae that contain endogenous juvenile hormone. In the last larval instar the juvenoid induces enormously large somatic growth and postpones pupal ecdysis although it does not increase the overall metabolic intensity beyond certain limits.The metabolic changes caused by the juvenoid were more pronounced when connected with formation of the supernumerary larval instars. After this developmental change the larvae further continued to grow on and turned finally into the conspicuous giants. The overall metabolic capacity of the supernumerary larval tissues surpassed the above limitations determined by the body of a normal last instar larva. However, unlike in some other species, there was no hypermetabolic response to juvenoid treatments in this material.     

Disruptive developmental effects of benzyl-1,3-benzodioxole derivatives on unparasitized and parasitized Manduca sexta larvae

Treatment of tobacco hornworm larvae with the benzyl-1,3-benzodioxole derivative J-2710 immediately after ecdysis to the fourth instar disrupted development either during the moult to the fifth instar or shortly thereafter. Larvae given topical applications of 100 μg J-2710 in 1 μl acetone suffered 100% mortality, often after secreting moulting fluid in large pockets between the epidermis and the cuticle later in the fourth instar. Larvae that successfully ecdysed had abnormalities of the mouthparts and cervix that interfered with normal feeding, inhibiting growth in the fifth instar. Larvae of the gregarious endoparasitic wasp Cotesia congregata (=Apanteles congregatus) frequently failed to emerge from host Manduca sexta larvae treated with high doses of J-2710, particularly when the host failed to feed normally. Less potent disruptive effects on Manduca and Cotesia were seen after treatment of larvae with the derivatives J-3370 and J-2581.No anti-juvenile hormone action of J-2710 was observed. J-2710-treated M. sexta larvae showed no precocious metamorphosis and the developmental effects of J-2710 were not prevented by co-application of the juvenile hormone analogue methoprene in doses ranging from 1 to 100 μg/larva. Moreover, J-2710 had no effect on the action of methoprene in the black larval assay for juvenile hormone-like activity, unlike results reported to occur using the Galleria wax wound assay.     

Locust solid cuticle—A time sequence of mechanical properties

Studies on locust femur solid cuticle indicate that there is a continuum of biomechanically important properties from one instar to the next, and that there is a constant ratio of stiffness to mass which is only interrupted at ecdysis. The biochemical nature of both larval and adult solid cuticle is interpreted in terms of both the tangent modulus and the breaking strength of the femur cuticle. The mechanical behaviour of the different layers in the cuticle is discussed.     

Exuviae eating: a nitrogen meal?

Many insects eat their cast cuticle (exuviae) after moulting. The functional significance of this behaviour has not been addressed experimentally. I tested the hypothesis that exuviae eating constitutes a meal, so the animal recycles its nitrogen content. Nitrogenous compounds (protein and chitin) are major components of the cuticle in Periplaneta americana, accounting for as much as 87% of the total weight. It was found that insects almost invariably ate their exuviae during their larval life. The frequency of the behaviour decreased in newly emerged adults and varied between the sexes, males eating their exuviae less frequently than females. This may be due to the extra nitrogen endowment which females need for reproduction. Aposymbiotic animals, which lack the supply of essential amino acids from endosymbiotic bacteria, always ate their exuviae regardless of sex. When animals were reared on different diets throughout their larval life protein level in the diet correlated with exuviae eating. Animals reared on a low protein diet showed the highest levels of exuviae eating; animals reared on a high protein diet showed the highest levels of exuviae rejection. Analysis of the frass produced after exuviae meals showed that over 58% of the nitrogen present in the exuviae was recycled. This demonstrated that cockroaches digested nitrogenous compounds contained in the cuticle. The possibility that the exuviae meal has other functions is discussed, although the evidence supports a nutritional role.     

Comparison between the sclerotization of adult and larval cuticle in Schistocerca gregaria

Femur cuticle from fifth instar larvae of the desert locust, Schistocerca gregaria, has been characterized with respect to composition, rate of deposition, and rate of sclerotization. The results are compared with those from adult cuticle of the same species. The protein compositions of the two types of cuticle are very similar, but the rates of deposition of both protein and chitin are different. The main difference is, however, that sclerotization is restricted to the first day after ecdysis in larval cuticle, whereas in adult cuticle sclerotization continues for at least a couple of weeks. The result is that the endocuticle remains untanned in the larvae, whereas in the adults the whole cuticle becomes tanned.     

Asian citrus psyllid stylet morphology and applicability to the model for inter-instar stylet replacement in the potato psyllid

In Hemiptera, presumptive stylets for each consecutive postembryonic instar are manufactured prior to ecdysis to replace the ecdysial stylets discarded with the exuviae. With the discovery that the bacterium “Candidatus” Liberibacter solanacearum accesses the tissues involved in the stylet replacement process of the potato psyllid, a hypothesis was formed that the bacterium could adhere to the stylets of freshly emerged instars and hence gain access to the host plant when feeding is resumed. Although unproven, it was imperative that a model for stylet replacement be built. Stylet morphology and the stylet replacement process of the Asian citrus psyllid (ACP), vector of “C.” L. asiaticus, causal pathogen of citrus greening disease, are comparable to the potato psyllid model system. Morphology consists of a basal terminus with its tab-shaped auricle, a base, shaft, and an apical terminus. Each of the four auricles act as a platform for the replacement apparatus, which is compacted into a tight aggregate of cells, the ‘end-cap’. As modeled, on apolysis of larval instar hypodermis, the aggregate ‘deconstructs’ and expands into a snail shell-shaped tube, the ‘atrium’, that houses the presumptive stylet as it is synthesized. Completed stylets then despool from the atrium and are fitted into their functional positions as the next instar emerges from its exuviae.     

Requirements for molting of the crochet epidermis of the tobacco hornworm larva in vivo and in vitro

Summary In the tobacco hornworm,Manduca sexta, the epidermis which underlies the larval crochets is the first tissue to become independent of the prothoracic glands (PG) in a larval molt. In each successive larval molt, crochet forming cells increase in size, form hooks at their distal ends and, finally, secrete cuticle. This paper examines the endocrine requirements for competence to molt and describes parallel cultures in vivo and in vitro to define the hormonal control of crochet molting. When implanted into a fourth instar host larva prior to initiation of the last larval molt, competent crochet epidermis molted, forming crochets synchronously with its host. In the fourth instar, competence to form crochets is attained slowly during the first two days following ecdysis from the third instar. During the feeding phase of the fifth (last) instar, the crochet epidermis remains competent to molt (to form an extra sixth instar set of crochets) until the larva attains a weight of about 4.5 gm. Then, concurrent with the decline in the titer of juvenile hormone (JH) in the hemolymph, competence to form crochets declines. A similar loss of competence did not occur when fourth instar crochet epidermis was exposed to a declining JH titer by culture in either fourth instar isolated abdomens for 72 h or in fifth instar host larvae between 4 and 7 gm. Responses of crochet epidermis cultured in vitro also were examined. Competent fourth instar crochet epidermis formed crochets following 3–6 h exposure to ecdysone in vitro. Six ×10^–7M -ecdysone was required for 50% response, whereas a 10–50-fold higher concentration of -ecdysone was necessary. Although formation of morphologically complete crochets in vitro proceeded with similar time course to that in situ, no molt-induced growth occurred in vitro. When crochet epidermis was exposed to ecdysone in vitro immediately after explantation, exogenous JH was not required for molting. But when tissue was first cultured for 72 h without hormones, subsequent molting in vitro could not be elicited, although molting still could occur when the tissue subsequently was implanted into a fourth instar host. Exposure to corpora allata or to JH during the 72 h of culture in vivo partially prevented the loss in capacity to respond to ecdysone in vitro, suggesting that JH may be one factor involved directly or indirectly in maintenance of tissue responsiveness.Preliminary presentation of some of this work given at the December, 1973 Meeting of the American Society of Zoologists (Fain and Riddiford, 1973)     

Action in vivo d'ecdysones sur la morphogénèse imaginale d'Aeshna cyanea (Odonata)

Single amounts of α or β ecdysone were injected during the last larval instar of Aeshna cyanea at various times after ecdysis. In these experimental conditions, α and β ecdysone had similar effects. Very large amounts of brown or black cuticle appeared on the tarsal claws soon after hormone injection, so that the cuticular synthesis of the larvae which were injected at the beginning of the last stage appears about two or three times more quickly than in controls. Nearly all the larval characters were exhibited by animals injected on the day of or the day after the last larval ecdysis. If the hormonal injection was further delayed, only adultoid forms were obtained. No perfect adults appeared. The effects evoked by α or β ecdysone may be different from one organ to another.On the other hand, some results were different according to the type of ecdysone. Darkening of the tarsal claws (and perhaps sclerotization) appears sooner when β ecdysone is supplied. The morphology of the external organs which degenerate during metamorphosis is not always the same after injection of equal amounts of α or β ecdysone at the same time. The regression of the larval organs seems to be more explicit and appears sooner when β ecdysone was administrated. The morphogenesis of the organs which grow during metamorphosis was either weaker or non-existent with β ecdysone.These results are discussed with regard to previous work.     

Developmental profiles of the mRNAs for Manduca arylphorin and two other storage proteins during the final larval instar of Manduca sexta

When fat body mRNA from the tobacco hornworm larva, Manduca sexta, was translated in a rabbit reticulocyte lysate system, three major polypeptides were found, each having a different developmental profile. One mRNA coded for a 74 kilodalton (K) polypeptide doublet precipitated by an antibody to the arylphorin (manducin). This mRNA was present only during the intermolt feeding phase of the penultimate and the final larval instars. Its appearance 16–24 hr after larval ecdysis was dependent upon the incoming nutrient supply and independent of the juvenile hormone (JH) level. Immunoblots of proteins of the fat body, epidermis, and cuticle revealed the presence of arylphorin in all three tissues. Additionally, several small polypeptides that cross-reacted with the arylphorin antibody were found in the fat body during and up to 24 hr after the last larval molt and in the tanning pupal cuticle. The larval epidermis was also found to contain a small amount of arylphorin mRNA. At the time of the JH decline prior to the onset of metamorphosis, a female-specific mRNA coding for a 79 K translation product appeared. In allatectomized larvae this mRNA was detectable earlier, and its appearance in intact larvae was prevented by application of methoprene, indicating that JH regulates its appearance. At wandering a new mRNA that also codes for a 79 K polypeptide appeared in both sexes and was the major messenger present during the prepupal stage. Neither it nor the female-specific mRNA were translatable after pupal ecdysis.     

Cuticular morphogenesis during continuous growth of the final instar larva of a moth

The larvae of the tobacco hornworm, Manduca sexta, grow continuously. During the feeding period of the fifth larval instar their weight increases ten-fold (ca. 1·2–12 g) accompanied by a four-fold expansion of the surface area of the abdominal cuticle. We have found that this cuticle contains structures which facilitate its expansion. Folds in the epicuticle (papillae) flatten as the cuticle expands. The endocuticle, in contrast, does not unfold but rather is plastically deformed. This plastic deformation is assisted by vertical structures in the cuticle (cuticular columns) which are more easily deformed than the surrounding lamellate cuticle. The head capsule cuticle, which does not expand as the larva grows, lacks papillae and cuticular columns. Thus, these are specialized structures that are reserved for cuticle that must expand as the larva grows.     

Effect of parasitism byAscogaster reticulatus [Hym.: Braconidae] on growth of the host,Adoxophyes sp. [Lep.: Tortricidae]

The developmental interaction between the egg/larval parasitoid,Ascogaster reticulatus Watanabe (Hymenoptera: Braconidae) and its host,Adoxophyes sp. (Lepidoptera: Tortricidae) was examined. Prior to the egress of a final-instar parasitoid larva from the 4^th-instar host larva, host weight decreased by 22% from the maximum weight. The final body weight of a host larva was 27% of the maximum weight of a healthy 5^th-instar host. Food consumption was significantly reduced in both 3^rd-and 4^th-instar parasitized larvae compared with healthy ones. In the 4^th instar, a parasitized larva consumed 28% less artificial diet and produced less frass than a healthy larva. The growth rate of the endoparasitoid larvae greatly increased after their host's molt to the 4^th instar. Parasitoid larval volume increased 40 fold in the 4^th-instar host.      

抑太保对亚洲玉米螟表皮酚氧化酶及几丁质酶活力的影响

采用饲喂法,抑太保(Atabron,IKI-7899)对亚洲玉米螟Ostrinia furnacalis幼虫的LC₅₀约是灭幼脲Ⅰ号的1/10.体内和体外实验表明,抑太保及灭幼脲Ⅰ号均能导致幼虫表皮酚氧化酶及几丁质酶活力显著增高.与对照相比,饲喂抑太保及灭幼脲Ⅰ号,五龄幼虫表皮酚氧化酶活力分别增长276.00%和28.00%、几丁质酶活力分别增长42.25%和13.62%;腹腔注射抑太保及灭幼脲Ⅰ号,五龄幼虫表皮酚氧化酶活力分别增长74.86%和37.98%、几丁质酶活力分别增长68.18%和28.86%.我们认为,酚氧化酶及几丁质酶活力的增高应是抑太保及灭幼脲Ⅰ号导致玉米螟死亡的重要原因.     

Dynamics of the pregnancy cycle in the tsetse Glossina morsitans

A normal pregnancy in tsetse involves the successful integration of larval development with maternal activity. At 25°C, ovulation in Glossina morsitans occurs 1 hr after the previous larviposition, the egg hatches on day 3·8 (1·57 mm length, 0·09 mg dry wt.), ecdysis to second instar occurs on day 4·9 (2·3 mm, 0·30 mg), the third instar cuticle is formed on day 6·8 (4·5 mm, 5·0 mg), and parturition occurs on day 9·0 (6·0 mm, 10·0 mg). Melanization of the in utero third instar follows a regular sequence over a 2 day period. Parturition follows a circadian pattern with a peak 9 hr after lights on (12 hr daily photophase). All instars receive nutriment from the female's milk gland. During early pregnancy the rate of milk synthesis is greater than rate of uptake by the larva, thus causing expansion of the secretory reservoirs. After day 6, the volume of the secretory reservoirs decreases, but as is indicated by nuclear volume and larval growth the rate of synthesis remains high until day 8. Feeding activity of the adult female is maximal on day 1, levels off at 60 per cent up to day 6, and then declines sharply towards the end of pregnancy. Oöcyte development proceeds in phase with larval development and thus minimizes a lag period between successive pregnancies.